U.S. patent number 6,919,345 [Application Number 10/477,446] was granted by the patent office on 2005-07-19 for sulfonate substituted pyrazol pyridine derivatives.
This patent grant is currently assigned to Bayer Aktiengesellschaft. Invention is credited to Cristina Alonso-Alija, Klaus Dembowsky, Achim Feurer, Dietmar Flubacher, Dieter Lang, Elisabeth Perzborn, Elke Stahl, Johannes-Peter Stasch, Alexander Straub, Stefan Weigand, Frank Wunder.
United States Patent |
6,919,345 |
Stasch , et al. |
July 19, 2005 |
Sulfonate substituted pyrazol pyridine derivatives
Abstract
This invention relates to pyrazol pyridine derivatives of
formula (I) ##STR1## wherein R.sup.1 is a radical of the formula
-O--SO.sub.2 --R.sup.3 in which R.sup.3 is optionally substituted
C.sub.1--6 --alkyl, optionally substituted C.sub.3--8 --cycloalkyl,
or optionally substituted phenyl; R.sup.2 is H, optionally
substituted C.sub.1--6 --alkyl --CO--or optionally substituted
C.sub.1--6 --alkyl--SO.sub.2 -- ; as well as salts, stereoisomers,
tautomers, and hydrates thereof. Pharmaceutical compositions
containing these materials and methods of using them in medical
treatment are also described and claimed.
Inventors: |
Stasch; Johannes-Peter
(Solingen, DE), Feurer; Achim (Wilhelmsfeld,
DE), Weigand; Stefan (Wuppertal, DE),
Stahl; Elke (Bergisch Gladbach, DE), Flubacher;
Dietmar (Freiburg, DE), Alonso-Alija; Cristina
(Haan, DE), Wunder; Frank (Wuppertal, DE),
Lang; Dieter (Velbert, DE), Dembowsky; Klaus
(Munich, DE), Straub; Alexander (Wuppertal,
DE), Perzborn; Elisabeth (Wuppertal, DE) |
Assignee: |
Bayer Aktiengesellschaft
(Leverkusen, DE)
|
Family
ID: |
7684395 |
Appl.
No.: |
10/477,446 |
Filed: |
April 22, 2004 |
PCT
Filed: |
April 30, 2002 |
PCT No.: |
PCT/EP02/04733 |
371(c)(1),(2),(4) Date: |
April 22, 2004 |
PCT
Pub. No.: |
WO02/09259 |
PCT
Pub. Date: |
November 21, 2002 |
Foreign Application Priority Data
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May 11, 2001 [DE] |
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101 22 894 |
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Current U.S.
Class: |
514/256; 544/327;
544/298; 544/328; 514/269 |
Current CPC
Class: |
A61P
9/00 (20180101); A61P 7/02 (20180101); A61P
43/00 (20180101); A61P 29/00 (20180101); C07D
471/04 (20130101); A61P 9/10 (20180101); A61P
15/10 (20180101); A61P 25/00 (20180101); A61P
9/12 (20180101) |
Current International
Class: |
C07D
471/04 (20060101); C07D 471/00 (20060101); C07D
471/04 (); A71K 031/4162 () |
Field of
Search: |
;544/298,327,328
;514/269,256 |
References Cited
[Referenced By]
U.S. Patent Documents
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6180656 |
January 2001 |
Furstner et al. |
6451805 |
September 2002 |
Straub et al. |
6462068 |
October 2002 |
Straub et al. |
6743798 |
June 2004 |
Straub et al. |
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Foreign Patent Documents
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19834045 |
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Feb 2000 |
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DE |
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19834047 |
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Feb 2000 |
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DE |
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10057751 |
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May 2002 |
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DE |
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9816223 |
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Apr 1998 |
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WO |
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9816507 |
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Apr 1998 |
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WO |
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9823619 |
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Jun 1998 |
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WO |
|
0006567 |
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Feb 2000 |
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WO |
|
0006568 |
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Feb 2000 |
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WO |
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0006569 |
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Feb 2000 |
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WO |
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0021954 |
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Apr 2000 |
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WO |
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Other References
Prandoni, The treatment of venous thromboembolic disorders: new
challenges and opportunities, Journal of Hematology, vol.
88(05):610-613, May 2003. .
Wolin et al., Oxidant-Nitric Oxide Signalling Mechanisms in
Vascular Tissue, Biochemistry (Moscow), vol. 63, No. 7, 810/958,
1998. .
Layzer, Degenerative Diseases of the Nervous System, Cecil Textbook
of Medicine, 20.sup.th Edition, vol. 2, pp. 2050-2057, 1996. .
Damasio, Alzheimer's Disease and Related Dementias, Cecil Textbook
of Medicine, 20.sup.th Edition, vol. 2, pp. 1992-1996, 1996. .
Fisker, PubMed Abstract (J. Endocrinol. Invest. 22(5 Suppl):89-93,
1999. .
Ko et al., YC-1, a Novel Activator of Platelet Guanylate Cyclase,
Blood, 84, 4226-4233 (1994). .
Mulsch, et al., Effect of YC-1, an NO-independent,
Superoxide-Sensitive Stimulator of Soluble Guanylyl Cyclase, on
Smooth Muscle Responsiveness to Nitrovasodilators, Brit. J. Pharm.
120, 681-689 (1997). .
Glass et al., Stimulation of Human Platelet Guanylate Cyclase by
Fatty Acids, J. Biol. Chem., 252, 1279-1285 (1977). .
Pettibone et al., A Structurally Novel Stimulator of Guanylate
Cyclase with Long-lasting Hypotensive Activity in the Dog, European
J. Pharm. 116, 307-312 (1985). .
Yu et al., Vasorelaxant Effect of Isoliquiritigenin, a Novel
Soluble Guanylate Cyclase Activator , in Rat Aorta, Brit. J. Pharm.
114, 1587-1594 (1995)..
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Primary Examiner: Rao; Deepak
Parent Case Text
This application is a 371 of PCT/EP02/04733 filed Apr. 30, 2002.
Claims
We claim:
1. A compound of the formula (I) ##STR26## in which R.sup.1 is a
radical of the formula --O--SO.sub.2 --R.sup.3, where R.sup.3 is a
radical from the group consisting of optionally substituted
C.sub.1-6 -alkyl, optionally substituted C.sub.3-8 -cycloalkyl, or
optionally substituted phenyl; R.sup.2 is H, optionally substituted
C.sub.1-6 -alkyl-CO or optionally substituted C.sub.1-6
-alkyl-SO.sub.2 --; or a salt, stereoisomer, tautomer or hydrate
thereof.
2. The compound as claimed in claim 1, in which R.sup.1 is a
radical of the formula --O--SO.sub.2 --R.sup.3, where R.sup.3 is a
radical selected from the group consisting of C.sub.1-6 -alkyl
which is optionally substituted by one to three halogen radicals,
and C.sub.3-8 -cycloalkyl; and R.sup.2 is H, C.sub.1-6 -alkyl-CO
which is optionally substituted by one to three halogen radicals,
or C.sub.1-6 -alkyl-SO.sub.2 -- which is optionally substituted by
one to three halogen radicals.
3. The compound as claimed in claim 1, in which R.sup.1 is a
radical of the formula --O--SO.sub.2 --R.sup.3, where R.sup.3 is a
radical selected from the group consisting of methyl, ethyl,
n-propyl, isopropyl, n-butyl, n-pentyl, 1,1,1-trifluoro-4-n-butyl,
chloromethyl and cyclopropyl; and R.sup.2 is H or CH.sub.3 CO.
4. A process for preparing compounds of the formula I, comprising
reacting a compound of the formula (II) ##STR27##
with a compound of the formula (III) ##STR28##
in an organic solvent in the presence of a base with heating and
subsequent conversion of the ether group in the free hydroxyl group
to give a compound of the formula (IV) ##STR29##
and subsequent reaction with a compound of the formula X--SO.sub.2
--R.sup.2 in which X is a leaving group which can be replaced by a
hydroxyl group; and R.sup.2 has the meaning indicated in claim 1;
in an organic solvent in the presence of a base with heating to
give a compound of the formula (I).
5. A pharmaceutical composition comprising at least one compound of
the formula (I) as claimed in claim 1 plus a pharmaceutically
acceptable carrier.
6. A pharmaceutical composition comprising at least one compound of
the formula (I) as claimed in claim 1 in combination with at least
one organic nitrate or NO donor.
7. A pharmaceutical composition comprising at least one compound of
the formula (I) as claimed in claim 1 in combination with at least
one compound which inhibits the breakdown of cyclic guanosine
monophosphate (cGMP).
8. A method for the treatment of hypertension comprising
administering an effective amount of a compound of general formula
(I) as claimed in claim 1.
9. A method for the treatment of sexual dysfunction comprising
administering an effective amount of a compound of general formula
(I) as claimed in claim 1.
10. The method as claimed in claim 8 or claim 9, where the compound
of the general formula (I) as claimed in claim 1 is employed in
combination with at least one organic nitrate or NO donor or in
combination with at least one compound which inhibits the breakdown
of cyclic guanosine monophosphate (cGMP).
Description
The present invention relates to novel chemical compounds which
stimulate soluble guanylate cyclase, to the preparation thereof and
to the use thereof as medicaments, in particular as medicaments for
the treatment of cardiovascular disorders.
One of the most important cellular transmission systems in
mammalian cells is cyclic guanosine monophosphate (cGMP). Together
with nitric oxide (NO), which is released from the endothelium and
transmits hormonal and mechanical signals, it forms the NO/cGMP
system. Guanylate cyclases catalyze the biosynthesis of cGMP from
guanosine triposphate (GTP). The representatives of this family
disclosed to date can be divided both according to structural
features and according to the type of ligands into two groups: the
particulate guanylate cyclases which can be stimulated by
natriuretic peptides, and the soluble guanylate cyclases which can
be stimulated by NO. The soluble guanylate cyclases consist of two
subunits and very probably contain one heme per heterodimer, which
is part of the regulatory site. The latter is of central importance
for the mechanism of activation. NO is able to bind to the iron
atom of heme and thus markedly increase the activity of the enzyme.
Heme-free preparations cannot, by contrast, be stimulated by NO. CO
is also able to attach to the central iron atom of heme, but the
stimulation by CO is distinctly less than that by NO.
Through the production of cGMP and the regulation, resulting
therefrom, of phosphodiesterases, ion channels and protein kinases,
guanylate cyclase plays a crucial part in various physiological
processes, in particular in the relaxation and proliferation of
smooth muscle cells, in platelet aggregation and adhesion and in
neuronal signal transmission, and in disorders caused by an
impairment of the aforementioned processes. Under
pathophysiological conditions, the NO/cGMP system may be
suppressed, which may lead for example to high blood pressure,
platelet activation, increased cellular proliferation, endothelial
dysfunction, atherosclerosis, angina pectoris, heart failure,
thromboses, stroke, and myocardial infarction.
A possible way of treating such disorders which is independent of
NO and aims at influencing the cGMP signal pathway in organisms is
a promising approach because of the high efficiency and few side
effects which are to be expected.
Compounds, such as organic nitrates, whose effect is based on NO
have to date been exclusively used for the therapeutic stimulation
of soluble guanylate cyclase. NO is produced by bioconversion and
activates soluble guanylate cyclase by attaching to the central
iron atom of heme. Besides the side effects, the development of
tolerance is one of the crucial disadvantages of this mode of
treatment.
Some substances which directly stimulate soluble guanylate cyclase,
i.e. without previous release of NO, have been described in recent
years, such as, for example,
3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1, Wu et al.,
Blood 84 (1994), 4226; Mulsch et al., Br. J. Pharmacol. 120 (1997),
681), fatty acids (Goldberg et al, J. Biol. Chem. 252 (1977),
1279), diphenyliodonium hexafluorophosphate (Pettibone et al., Eur.
J. Pharmacol. 116 (1985), 307), isoliquiritigenin (Yu et al., Brit.
J. Pharmacol. 114 (1995), 1587) and various substituted pyrazole
derivatives (WO 98/16223).
In addition, WO 98/16507, WO 98/23619, WO 00/06567, WO 00/06568, WO
00/06569 and WO 00/21954 describe pyrazolopyridine derivatives as
stimulators of soluble guanylate cyclase. Also described in these
patent applications are pyrazolopyridines having a pyrimidine
residue in position 3. Compounds of this type have very high in
vitro activity in relation to stimulating soluble guanylate
cyclase. However, it has emerged that these compounds have some
disadvantages in respect of their in vivo properties such as, for
example, their behavior in the liver, their pharmacokinetic
behavior, their dose-response relation or their metabolic
pathway.
It was therefore the object of the present invention to provide
further pyrazolopyridine derivatives which act as stimulators of
soluble guanylate cyclase but do not have the disadvantages,
detailed above, of the compounds from the prior art.
This object is achieved according to the present inventions by the
compounds as claimed in claim 1. These novel pyrazolopyridine
derivatives are distinguished by a pyrimidine residue in position
3, which has a particular substitution pattern, namely a sulfonate
residue in position 5 of the pyrimidine ring and an amino group in
position 4 of the pyrimidine ring.
The present invention specifically relates to compounds of the
formula (I) ##STR2##
in which R.sup.1 is a radical of the formula --O--SO.sub.2
--R.sup.3, where R.sup.3 is a radical from the group consisting of
optionally substituted C.sub.1-6 -alkyl, optionally substituted
C.sub.3-8 -cycloalkyl, or optionally substituted phenyl; R.sup.2 is
H, optionally substituted C.sub.1-6 -alkyl-CO or optionally
substituted C.sub.1-6 -alkyl-SO.sub.2 --;
and salts, isomers and hydrates thereof.
Preference is given according to the present invention to compounds
of the formula (I) in which R.sup.1 is a radical of the formula
--O--SO.sub.2 --R.sup.3, where R.sup.3 is a radical from the group
consisting of C.sub.1-6 -alkyl which is optionally substituted by
one to three halogen radicals, or C.sub.3-8 -cycloalkyl; R.sup.2 is
H, C.sub.1-6 -alkyl-CO which is optionally substituted by one to
three halogen radicals, or C.sub.1-6 -alkyl-SO.sub.2 -- which is
optionally substituted by one to three halogen radicals;
and salts, isomers and hydrates thereof.
Particular preference is given in this connection to compounds of
the formula (I) in which R.sup.1 is a radical of the formula
--O--SO.sub.2 --R.sup.3, where R.sup.3 is a radical from the group
consisting of methyl, ethyl, n-propyl, isopropyl, n-butyl,
n-pentyl, 1,1,1-trifluoro-4-n-butyl, chloromethyl or cyclopropyl;
R.sup.2 is H or CH.sub.3 CO;
and salts, isomers and hydrates thereof.
The compounds of the invention of the general formula (I) may also
exist in the form of their salts. Salts which may generally be
mentioned here are those with organic or inorganic bases or
acids.
For the purposes of the present invention, physiologically
acceptable salts are preferred. Physiologically acceptable salts of
the compounds of the invention may be salts of the substances of
the invention with mineral acids, carboxylic acids or sulfonic
acids. Particularly preferred examples are salts with hydrochloric
acid, hydrobromic acid, sulfuric acid, phosphoric acid,
methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid,
benzenesulfonic acid, naphthalenedisulfonic acid, acetic acid,
propionic acid, lactic acid, tartaric acid, citric acid, fumaric
acid, maleic acid or benzoic acid.
Physiologically acceptable salts may likewise be metal or ammonium
salts of the compounds of the invention which have a free carboxyl
group. Particularly preferred examples are sodium, potassium,
magnesium or calcium salts, and ammonium salts derived from ammonia
or organic amines such as, for example, ethylamine, di- or
triethylamine, di- or triethanolamine, dicyclohexylamine,
dimethylaminoethanol, arginine, lysine or ethylenediamine.
The compounds of the invention may exist in stereoisomeric forms
which either are related as image and mirror image (enantiomers) or
which are not related as image and mirror image (diastereomers).
The invention relates both to the enantiomers or diastereomers and
to the mixtures thereof in each case. The racemic forms can, just
like the diastereomers, be separated in a known manner, for example
by chromatographic separation, into the stereoisomerically pure
constituents. Double bonds present in the compounds of the
invention may be in the cis or trans configuration (Z or E
form).
Certain compounds may moreover exist in tautomeric forms. This is
known to the skilled worker, and the scope of the invention
likewise covers such compounds.
The compounds of the invention may additionally occur in the form
of their hydrates, where the number of water molecules bound to the
molecule depends on the particular compound of the invention.
Unless otherwise indicated, for the purposes of the present
invention the substituents generally have the following
meaning:
Alkyl is generally a straight-chain or branched hydrocarbon radical
having 1 to 6 carbon atoms. Examples which may be mentioned are
methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl,
isopentyl, hexyl, isohexyl.
Cycloalkyl is generally a cyclic hydrocarbon radical having 3 to 8
carbon atoms. Cyclopropyl, cyclopentyl and cyclohexyl are
preferred. Examples which may be mentioned are cyclopentyl,
cyclohexyl, cycloheptyl and cyclooctyl.
Halogen is for the purposes of the invention fluorine, chlorine,
bromine and iodine.
The compounds of the invention of the formula (I) can be prepared
by reacting the compound of the formula (II) ##STR3##
with compounds of the formula (III) ##STR4##
in an organic solvent in the presence of a base with heating and
subsequent conversion of the ether group into the free hydroxyl
group to compounds of the formula (IV) ##STR5##
and subsequent reaction with compounds of the formula X--SO.sub.2
--R.sup.2 in which X is a leaving group which can be replaced by a
hydroxyl group; R.sup.2 has the meaning indicated above;
in an organic solvent in the presence of a base with heating to
give compounds of the formula (I).
The compound of the formula (II) can be prepared as shown in the
following reaction scheme: ##STR6##
The compound of the formula (II) can be obtained in a multistage
synthesis from the sodium salt of ethyl cyanopyruvate which is
known from the literature (Borsche and Manteuffel, Liebigs. Ann.
Chem. 1934, 512, 97). Reaction thereof with 2-fluorobenzylhydrazine
with heating and under a protective gas atmosphere in an inert
solvent such as dioxane results in ethyl
5-amino-1-(2-fluorobenzyl)pyrazole-3-carboxylate, which cyclizes to
the corresponding pyridine derivative by reaction with
dimethylaminoacrolein in acidic medium under a protective gas
atmosphere and with heating. This pyridine derivative ethyl
1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboxylate is
converted by a multistage sequence consisting of conversion of the
ester with ammonia into the corresponding amide, dehydration with a
dehydrating agent such as trifluoroacetic anhydride to give the
corresponding nitrile derivative, reaction of the nitrile
derivative with sodium ethoxide and final reaction with ammonium
chloride into the compound of the formula (II).
The compound of the formula (III) can be prepared from the
compounds, which can be purchased (e.g. from Aldrich),
t-butoxybis(dimethylamino)methane and methoxyacetonitrile by
reacting these reactants preferably in equimolar amounts preferably
under atmospheric pressure and stirring the reaction solution for
several hours, for example 12 hours, at elevated temperature, for
example 60-110.degree. C., preferably 70-90.degree. C., in
particular 80.degree. C.
The reaction of the compounds of the formulae (II) and (III) to
give the compound of the formula (IV) can be carried out by
employing the reactants in equimolar amounts or by using the
compound of the formula (III) in slight excess in an organic
solvent, for example an alcohol, preferably isoamyl alcohol, in the
presence of a small amount of a base, for example an organic amine,
in particular piperidine, preferably under atmospheric pressure and
stirring the reaction solution for several hours, for example 12
hours, at elevated temperature, for example 60-130.degree. C.,
preferably 80-120.degree. C., in particular 110.degree. C., and
subsequent liberation of the hydroxyl group by reacting the
compound obtained in this way with a preferably equimolar amount of
a thiol such as, for example, thiophenol in the presence of a small
amount of a base such as an alkali metal base, for example an
alkali metal carbonate, preferably potassium carbonate, in an
organic solvent such as, for example, 1-methyl-2-pyrrolidone,
preferably under atmospheric pressure and stirring the reaction
solution for several hours, for example 1 hour, at elevated
temperature, for example 100-200.degree. C., preferably
150-200.degree. C.
The compound of the formula (IV) obtained in this way can be
converted into the compounds of the formula (I) of the invention by
reaction with an equimolar amount or of a slight excess of a
sulfonyl compound of the formula XSO.sub.2 R.sub.2. The reaction is
carried out in the presence of a small amount of a base such as an
organic amine, preferably pyridine, preferably under atmospheric
pressure and stirring the reaction solution for several hours, for
example 12 hours, at elevated temperature, for example
40-80.degree. C., preferably 50-70.degree. C. The sulfonyl
compounds can be purchased or obtained in a manner known to the
skilled worker.
The compounds of the invention of the general formula (I) show a
valuable range of pharmacological effects which could not be
predicted.
The compounds according to the invention of the general formula (I)
bring about vasorelaxation and an inhibition of platelet
aggregation and lead to a reduction in blood pressure and an
increase in coronary blood flow. These effects are mediated by
direct stimulation of soluble guanylate cyclase and an
intracellular increase in cGMP. In addition, the compounds
according to the invention of the general formula (I) enhance the
effect of substances which increase the cGMP level, such as, for
example, EDRF (endothelium derived relaxing factor), NO donors,
protoporphyrin IX, arachidonic acid or phenylhydrazine
derivatives.
They can therefore be employed in medicaments for the treatment of
cardiovascular disorders such as, for example, for the treatment of
high blood pressure and heart failure, stable and unstable angina
pectoris, peripheral and cardiac vascular disorders, of
arrhythmias, for the treatment of thromboembolic disorders and
ischemias such as myocardial infarction, stroke, transistorily and
ischemic attacks, disturbances of peripheral blood flow, prevention
of restenoses as after thrombolysis therapies, percutaneously
transluminal angioplasties (PTAs), percutaneously transluminal
coronary angioplasties (PTCAs), bypass and for the treatment of
arteriosclerosis, asthmatic disorders and diseases of the
urogenital system such as, for example, prostate hypertrophy,
erectile dysfunction, female sexual dysfunction, osteoporosis,
gastroparesis and incontinence.
The compounds described in the present invention of the general
formula (I) also represent active ingredients for controlling
central nervous system diseases characterized by disturbances of
the NO/cGMP system. They are suitable in particular for improving
perception, concentration, learning or memory after cognitive
impairments like those occurring in particular in association with
situations/diseases/syndromes such as mild cognitive impairment,
age-associated learning and memory impairments, age-associated
memory loss, vascular dementia, craniocerebral trauma, stroke,
dementia occurring after strokes (post stroke dementia),
post-traumatic craniocerebral trauma, general concentration
impairments, concentration impairments in children with learning
and memory problems, Alzheimer's disease, vascular dementia, Lewy
body dementia, dementia with degeneration of the frontal lobes
including Pick's syndrome, Parkinson's disease, progressive nuclear
palsy, dementia with corticobasal degeneration, amyolateral
sclerosis (ALS), Huntington's disease, multiple sclerosis, thalamic
degeneration, Creutzfeld-Jacob dementia, HIV dementia,
schizophrenia with dementia or Korsakoff's psychosis. They are also
suitable for the treatment of central nervous system disorders such
as states of anxiety, tension and depression, CNS-related sexual
dysfunctions and sleep disturbances, and for controlling
pathological disturbances of the intake of food, stimulants and
addictive substances.
The active ingredients are furthermore also suitable for
controlling cerebral blood flow and thus represent effective agents
for controlling migraine.
They are also suitable for the prophylaxis and control of the
sequelae of cerebral infarctions such as stroke, cerebral ischemias
and craniocerebral trauma. The compounds of the invention of the
general formula (I) can likewise be employed for controlling states
of pain.
In addition, the compounds of the invention have an
anti-inflammatory effect and can therefore be employed as
anti-inflammatory agents.
Furthermore, the invention encompasses the combination of the
compounds of the invention of the general formula (I) with organic
nitrates and NO donors.
Organic nitrates and NO donors for the purposes of the invention
are generally substances which display their therapeutic effect via
release of NO or NO species. Preference is given to sodium
nitroprusside, nitroglycerine, isosorbide dinitrate, isosorbide
mononitrate, molsidomine and SIN-1.
In addition, the invention encompasses the combination with
compounds which inhibit breakdown of cyclic guanosine monophosphate
(cGMP). These are in particular inhibitors of phosphodiesterases 1,
2 and 5; nomenclature of Beavo and Reifsnyder (1990), TiPS 11 pp.
150 to 155. These inhibitors potentiate the effect of the compound
of the invention, and the desired pharmacological effect is
increased.
BIOLOGICAL INVESTIGATIONS
Vasorelaxant Effect in vitro
Rabbits are stunned by a blow to the back of the neck and are
exsanguinated. The aorta is removed, freed of adherent tissue,
divided into rings 1.5 mm wide and put singly under tension in 5 ml
organ baths containing carbogen-gassed Krebs-Henseleit solution at
37.degree. C. with the following composition (mM): NaCl: 119; KCl:
4.8; CaCl.sub.2.times.2 H.sub.2 O: 1; MgSO.sub.4.times.7 H.sub.2 O;
1.4; KH.sub.2 PO.sub.4 : 1.2; NaHCO.sub.3 : 25; glucose: 10. The
force of contraction is detected with Statham UC2 cells, amplified
and digitized via A/D converters (DAS-1802 HC, Keithley Instruments
Munich) and recorded in parallel on chart recorders. A contraction
is generated by adding phenylephrine to the bath cumulatively in
increasing concentration. After several control cycles, the
substance to be investigated is investigated in each further run in
increasing dosage in each case, and the height of the contraction
is compared with the height of the contraction reached in the last
preceding run. The concentration necessary to reduce the height of
the control value by 50% (IC.sub.50) is calculated from this. The
standard application volume is 5 .mu.l, and the DMSO content in the
bath solution corresponds to 0.1%. The results are listed in table
1 below:
TABLE 1 Vasorelaxant effect in vitro Example No. IC.sub.50 [nM] 1
700 2 580 3 300 4 710 5 520 7 440 10 2020
Determination of the Liver Clearance in vitro
Rats are anesthetized, heparinized, and the liver is perfused in
situ via the portal vein. Primary rat hepatocytes are then obtained
ex vivo from the liver using collagenase solution.
2.multidot.10.sup.6 hepatocytes per ml were incubated at 37.degree.
C. with the same concentration in each case of the compound to be
investigated. The decrease of the substrate to be investigated over
time was determined bioanalytically (HPLC/UV, HPLC/fluorescence or
LC/MSMS) at 5 points in time in each case in the period from 0-15
min after the start of incubation. From this, the clearance was
calculated by means of the cell count and liver weight.
Determination of the Plasma Clearance in vivo
The substance to be investigated is administered as a solution
intravenously to rats via the tail vein. At fixed points in time,
blood is taken from the rats, heparinized and plasma is obtained
therefrom by conventional measures. The substance is quantified
bioanalytically in the plasma. The pharmacokinetic parameters are
calculated from the plasma concentration-time courses determined in
this way by means of conventional non-compartmental methods used
for this purpose.
The present invention includes pharmaceutical preparations which,
besides nontoxic, inert pharmaceutically suitable carriers,
comprises the compounds of the invention of the general formula
(I), and processes for producing these preparations.
The active ingredient may be present where appropriate in one or
more of the carriers indicated above also in microencapsulated
form.
The therapeutically effective compounds of the general formula (I)
ought to be present in the pharmaceutical preparations mentioned
above in a concentration of about 0.1 to 99.5, preferably of about
0.5 to 95, % by weight of the complete mixture.
The pharmaceutical preparations mentioned above may, apart from the
compounds of the invention of the general formula (I), also
comprise other active pharmaceutical ingredients.
It has generally proved advantageous both in human and in
veterinary medicine to administer the active ingredient(s) of the
invention in total amounts of about 0.01 to about 700, preferably
0.01 to 100, mg/kg of body weight per 24 hours, where appropriate
in the form of a plurality of single doses, to achieve the desired
results. A single dose comprises the active ingredient(s) of the
invention preferably in amounts of about 0.1 to about 80, in
particular 0.1 to 30, mg/kg of body weight.
The present invention is described in more detail below by means of
nonrestrictive preferred examples. Unless indicated elsewhere, all
quantitative data relate to percentages by weight.
EXAMPLES
Abbreviations
RT: room temperature EA: ethyl acetate MCPBA: m-chloroperoxybenzoic
acid BABA: n-butyl acetate/n-butanol/glacial acetic acid/phosphate
buffer pH 6 (50:9:25.25; org. phase) DMF: N,N-dimethylformamide
Mobile Phases for the Thin-layer Chromatography
T1 E1: toluene-ethyl acetate (1:1) T1 EtOH1: toluene-methanol (1:1)
C1 E1: cyclohexane-ethyl acetate (1:1) C1 E2: cyclohexane-ethyl
acetate (1:2)
Methods for Establishing the HPLC Retention Times
Method A (HPLC-MS):
Eluent: A = CH.sub.3 CN B = 0.6 g 30% HCl/l H.sub.2 O Flow rate:
0.6 ml/min Column oven: 50.degree. C. Column: symmetry C18 2.1 *
150 mm
Gradient:
Time (min) % A % B Flow rate (ml/min) 0 10 90 0.6 4 90 10 0.6 9 90
10 0.8
Method B (HPLC):
Eluent: A = 5 ml HClO.sub.4 /l H.sub.2 O, B = CH.sub.3 CN Flow
rate: 0.75 ml/min L-R temperature: 30.00.degree. C. 29.99.degree.
C. Column: Kromasil C18 60 * 2 mm
Gradient:
Time (min) % A % B 0.50 98 2 4.50 10 90 6.50 10 90 6.70 98 2 7.50
98 2
Method C (HPLC):
Eluent: A = H.sub.3 PO.sub.4 0.01 mol/l, B = CH.sub.3 CN Flow rate:
0.75 ml/min L-R temperature: 30.01.degree. C. 29.98.degree. C.
Column: Kromasil C18 60 * 2 mm
Gradient:
Time (min) % A % B 0.00 90 10 0.50 90 10 4.50 10 90 8.00 10 90 8.50
90 10 10.00 90 10
Method D (chiral HPLC):
Eluent: 50% isohexane, 50% ethanol Flow rate: 1.00 ml/min
Temperature: 40.degree. C. Column: 250 * 4.6 mm, packed with
Chiralcel OD, 10 .mu.m
Method E (HPLC-MS):
Eluent: A = CH.sub.3 CN B = 0.3 g 30% HCl/l H.sub.2 O Flow rate:
0.9 ml/min Column oven: 50.degree. C. Column: Symmetry C18 2.1*150
mm
Gradient:
Time (min) % A % B Flow rate (ml/min) 0 10 90 0.9 3 90 10 1.2 6 90
10 1.2
STARTING COMPOUNDS
I. Synthesis of 3,3-bis(dimethylamino)-2-methoxypropionitrile
##STR7##
40.0 g (229.5 mmol) of ter-butoxybis(dimethylamino)methane and 16.3
g (229.5 mmol) of methoxyacetonitrile are stirred at 80.degree. C.
overnight. For working up, volatile material is stripped off in a
rotary evaporator, and the residue is distilled under high vacuum
in a Kugelrohr at 140.degree. C. The product contains, according to
the NMR spectrum (300 MHz, D.sub.6 -DMSO) the enamine as E/Z
mixture produced by elimination of dimethylamine. The product
mixture is employed without further purification in the next
reaction.
Yield: 24.7 g (60%).
II. Synthesis of
1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboxamidine
2A) Ethyl 5-amino-1-(2-fluorobenzyl)pyrazole-3-carboxylate
##STR8##
111.75 g (75 ml, 0.98 mol) of trifluoroacetic acid are added to 100
g (0.613 mol) of the sodium salt of ethyl cyanopyruvate (prepared
in analogy to Borsche and Manteuffel, Liebigs Ann. 1934, 512, 97)
while stirring efficiently in 2.5 l of dioxane at room temperature
under argon, and the mixture is stirred for 10 min, during which
most of the precursor dissolves. Then 85.93 g (0.613 mol) of
2-fluorobenzylhydrazine are added, and the mixture is boiled
overnight. After cooling, the sodium trifluoroacetate crystals
which have separated out are filtered off with suction and washed
with dioxane, and the crude solution is reacted further.
2B) Ethyl
1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboxylate
##STR9##
The solution obtained from 2 A) is mixed with 61.25 ml (60.77 g,
0.613 mol) of dimethylaminoacrolein and 56.28 ml (83.88 g, 0.736
mol) of trifluoroacetic acid and boiled under argon for 3 days. The
solvent is then evaporated in vacuo, and the residue is poured into
2 l of water and extracted three times with 1 l of ethyl acetate
each time. The combined organic phases are dried with magnesium
sulfate and concentrated in a rotary evaporator. Chromatography is
carried out on 2.5 kg of silica gel, eluting with a
toluene/toluene-ethyl acetate=4:1 gradient. Yield: 91.6 g (49.9% of
theory over two stages).
Melting point 85.degree. C.
R.sub.f (SiO.sub.2, T1E1): 0.83.
2C) 1-(2-Fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboxamide
##STR10##
10.18 g (34 mmol) of the ester obtained in example 2 B) are
introduced into 150 ml of methanol saturated with ammonia at
0-10.degree. C. Stirring at room temperature for two days is
followed by concentration in vacuo.
R.sub.f (SiO.sub.2,T1E1): 0.33.
2D) 3-Cyano-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine
##STR11##
36.1 g (133 mmol) of
1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboxamide from
example 2 C) are dissolved in 330 ml of THF, and 27 g (341 mmol) of
pyridine are added. Then, over the course of 10 min, 47.76 ml
(71.66 g, 341 mmol) of trifluoroacetic anhydride are added, during
which the temperature rises to 40.degree. C. The mixture is stirred
at room temperature overnight. The mixture is then poured into 1l
of water and extracted three times with 0.5l of ethyl acetate each
time. The organic phase is washed with saturated sodium bicarbonate
solution and with 1 N HCl, dried with MgSO4 and concentrated in a
rotary evaporator.
Yield: 33.7 g (100% of theory).
Melting point: 81.degree. C.
R.sub.f (SiO.sub.2, T1E1): 0.74.
2E) Methyl
(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboximidate
##STR12##
30.37 g (562 mmol) of sodium methoxide are dissolved in 1.5l of
methanol, and 36.45 g (144.5 mmol) of
3-cyano-1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine (from example
2 D) are added. The solution obtained after stirring at room
temperature for 2 hours is employed directly for the next
stage.
2F) 1-(2-Fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboxamidine
##STR13##
The solution of methyl
(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboximidate in
methanol obtained from example 2 E) is mixed with 33.76 g (32.19
ml, 562 mmol) of glacial acetic acid and 9.28 g (173 mmol) of
ammonium chloride and stirred under reflux overnight. The solvent
is evaporated in vacuo, the residue is thoroughly triturated with
acetone, and the precipitated solid is filtered off with
suction.
.sup.1 H-NMR (d.sub.6 -DMSO, 200 MHz): .delta.=5.93 (s, 2H);
7.1-7.5 (m, 4H); 7.55 (dd, 1H); 8.12 (dd, 1H); 8.30 (dd, 1H); 9.5
(bs, 4H exchangeable) ppm.
MS (EI): m/z=270.2 (M-HCl).
III. Synthesis of
2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]-5-methoxy-4-pyrimidi
nylamine
##STR14##
46.8 g (134.8 mmol) of
1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridine-3-carboximidamide
from example II are dissolved in isoamyl alcohol. To this are added
24.7 g (144.2 mmol) of
3,3-bis(dimethylamino)-2-methoxypropionitrile from example I and
1.15 g (1.33 ml, 13.5 mmol) of piperidine, and the mixture is left
to stir at 110.degree. C. for 3 days. For working up, it is cooled
to 0.degree. C., and the precipitated product is filtered off with
suction, washed thoroughly with cold diethyl ether and dried in a
vacuum oven at 50.degree. C.
Yield: 25.4 g (52.7%).
R.sub.f : 0.34 (dichloromethane/methanol 20:1).
.sup.1 H-NMR: (400 MHz, d.sub.6 -DMSO), .delta.=3.89 (2, 3H,
OCH.sub.3), 5.79 (s, 2H, CH.sub.2), 6.93 (br. s, 2H, NH.sub.2),
7.10-7.26 (m, 3H, Ar--H), 7.31-7.39 (m, 2H, Ar--H), 7.98 (s, 1H,
pyrimidine-H), 8.61 (dd, 1H, pyridine-H), 8.92 (dd, 1H,
pyridine-H).
MS: (ESI pos.), m/z=350.9 ([M+H].sup.+), 700.8 ([2M+H].sup.+).
IV. Synthesis of
4-amino-2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]-5-pyrimidino
l
##STR15##
25.3 g (72.2 mmol) of
2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]-5-methoxy-4-pyrimidi
nylamine from example III are dissolved in 500 ml of
1-methyl-2-pyrrolidone. To this are added 7.96 g (7.42 ml, 72.2
mmol) of thiophenol and 2.50 g (18.1 mmol) of potassium carbonate,
and the mixture is left to stir at 190.degree. C. for about 1 h.
For working up, the solvent is condensed off, and the residue is
mixed with half-conc. ammonium chloride solution and extracted
three times with ethyl acetate. Most of the product precipitates
during this. It is filtered off with suction and dried in a vacuum
oven at 50.degree. C.
Yield: 18.1 g (72.3%).
R.sub.f : 0.44 (dichloromethane/methanol 10:1).
.sup.1 H-NMR: (300 MHz, D.sub.6 -DMSO), .delta.=5.78 (s, 2H,
CH.sub.2), 6.66 (br. s, 2H, NH.sub.2), 7.09-7.38 (m, 5H, Ar--H),
7.82 (s, 1H, pyrimidine H), 8.60 (dd, 1H, pyridine H), 8.92 (dd,
1H, pyridine H), 9.4-10.2 (br. s, 1H, OH).
MS: (ESI pos.), m/z=337.3 ([M+H].sup.+), 673.3 ([2M+H].sup.+).
EXAMPLES
1.
4-Amino-2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]-5-pyrimidiny
l chloromethanesulfonate
##STR16##
400 mg (1.19 mmol) of
4-amino-2-[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]5-pyrimidinol
from example IV were suspended in 8.0 ml of pyridine, and 186.1 mg
(1.25 mmol) of chloromethanesulfonyl chloride were added. The
suspension was stirred at 60.degree. C. overnight and then water
was added to the mixture. The resulting precipitate was filtered
off with suction, washed several times with water and dried under
high vacuum.
Yield: 480 mg (77.3%).
.sup.1 H-NMR: (400 MHz, D.sub.6 -DMSO, .delta.=5.76 (s, 2H,
CH.sub.2), 5.82 (s, 2H, CH.sub.2), 6.66 (br. s, 2H, NH.sub.2),
7.10-7.26 (m, 3H, Ar--H), 7.30-7.42 (m, 2H, Ar--H), 7.59 (br. s,
2H, NH.sub.2), 8.31 (s, 1H, pyrimidine H), 8.65 (dd, 1H, pyridine
H), 8.93 (dd, 1H, pyridine H).
MS: (ESI pos.), m/z=449 ([M+H].sup.+), 897 ([2M+H].sup.+).
The following were prepared in an analogous manner:
Example Formula Yield (%) 1H-NMR 2 (from IV and methyl- sulfonyl
chloride) ##STR17## 89 (300 MHz, CDCl.sub.3): .delta.= 3.51(s, 3H),
5.83(s, 2H), 7.06-7.28(m, 3H), 7.31- 7.45(m, 2H), 7.58(bs, 2H),
8.28(s, 1H), 8.65(dd, J= 4.5 Hz, J=1.5 Hz, 1H), 8.94(dd, j=8.1 Hz,
J=1.5 Hz, 1H). 3 (from IV and ethyl- sulfonyl chloride) ##STR18##
89 (300 MHz, DMSO-d.sub.6, .delta. = 1.37(t, J=7.4 Hz, 3H), 3.71(q,
J=7.4 Hz, 2H), 5.83(s, 2H), 7.03-7.28 (m, 3H), 7.29-7.44(m, 2H),
7.52(bs, 2H), 8.29(s, 1H), 8.65(dd, J=4.5 Hz, J= 1.3 Hz, 1H),
8.93(dd, J= 8.1 Hz, J=1.3 Hz, 1H). 4 (from IV and cyclo- propyl-
sulfonyl chloride) ##STR19## 90 (300 MHz, DMSO-d.sub.6): d=
1.03-1.12(m, 2H), 1.14- 1.24(m, 2H), 3.21-3.39 (m, 1H), 5.83(s,
2H), 7.05- 7.28(m, 2H), 7.30-7.44 (m, 2H), 7.57(bs, 2H), 8.29 (s,
1H), 8.65(dd, J=4.5 Hz, J=1.3 Hz, 1H), 8-64 # (dd, J=7.9 Hz, J=1.3
Hz, 1H). 5 (from IV and isopropyl- sulfonyl chloride) ##STR20## 83
(300 MHz, DMSO-d.sub.6): d= 1.45(d, J=6.8 Hz, 6H), 4.04(sept, J=6.8
Hz, 1H), 5.83(s, 2H), 7.01-7.28 (m, 3H), 7.29-7.59(m, 4H), 8.29(s,
1H), 8.65(dd, J=4.4 Hz, J=1.5 Hz, 1H), 8.94(dd, J=8.1 Hz, J=1.5 Hz,
1H). 6 (from IV and n-pentyl- sulfonyl chloride) ##STR21## 32 (300
MHz, DMSO-d.sub.6): d= 0.87(t, J=7.0 Hz, 3H), 1.23-1.47=(m, 4h),
1.81 (quint, J=7.6 Hz, 2H), 3.70(t, J=7.7 Hz, 2H), 5.83(s, 2H),
7.05-7.27 (m, 3H), 7.31-7.46(m, 2H), 7.53(bs, 2H), 8.28(s, # 2H),
8.65(dd, J=4.4 Hz, J= 1.5 Hz, 1H), 8.93(dd, J= 8.1 Hz, J=1.5 Hz,
1H). 7 (from IV and 1,1,1- trifluoro- 4-butyl- sulfonyl chloride)
##STR22## 69 (300 MHz, DMSO-d.sub.6): d= 2.05(quint, J=7.9 Hz, 2H),
2.35-2.59(m, 2H), 3.83(t, J=7.7 Hz, 2H), 5.83(s, 2H), 7.06-7.28 (m,
3H), 7.29-7.44(m, 2H), 7.59(bs, 2H), 8.30(s, # 2H), 8.65(dd, J=4.5
Hz, J= 1.5 Hz, 1H), 8.93(dd, J= 8.1 Hz, J=1.5 Hz, 1H). 8 (from IV
and n-butyl- sulfonyl chloride) ##STR23## 99 (300 MHz,
DMSO-d.sub.6): d= 0.91(t, J=7.4Hz, 3H), 1.44(sex, J=7.4 Hz, 2H),
1.79(quint, J=7.4 Hz, 2H), 3.71(t, J=7.5 Hz, 2H), 5.83(s, 2H),
7.08- 7.27(m, 3H), 7.31-7.43 (m, 2H), 7.53(bs, 2H), 8.28 (s, 1H),
8.65(dd, J=4.5 # Hz, J=1.7 Hz, 1H), 8.93 (dd, J=8.1 Hz, J=1.7 Hz,
1H). 9 (from IV and n-propyl- sulfonyl chloride) ##STR24## 98 (300
MHz, DMSO-d.sub.6): d= 1.02(t, J=7.4 Hz, 3H), 1.84(sex, J=7.4 Hz,
2H), 3.68(t, J=7.6 Hz, 2H), 5.83(s, 2H), 7.08- 7.27 (m, 3H),
7.31-7.43(m, 2H), 7.53(bs, 2H), 8.28(s, 1H), 8.65(dd, J=4.5 Hz,
J=1.7 Hz, 1H), 8.93(dd, J= 8.1 Hz, J=1.7 Hz, 1H). 10 (from IV and
phenyl- sulfonyl chloride) ##STR25## 35.4 (300 MHz, DMSO-d.sub.6):
.delta. = 5.80(s, 2H), 7.08-7.24 (m, 7H), 7.66(t, J=7.6 Hz, 2H),
7.78-7.87(t, J=7.5 Hz, 1H), 7.97-8.03(m, 2H), 8.06(s, 1H), 8.64(dd,
J=4.3 Hz, J=1.5 Hz, 1H), 8.88(dd, J=8.1 Hz, J=1.7 Hz, 1H).
* * * * *